Serodiagnosis of Mycobacterium bovis infection in badgers: development of an indirect ELISA using a 25 kDa antigen

Goodger, Jane; Nolan, Ann; Russell, W. J.; Dalley, DJ; Thorns, C. J.; Stuart, FA; Croston, P; Newell, DG

doi:10.1136/vr.135.4.82

Cited by 87 publications

(65 citation statements)

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“…These results broadly agree with RT performance in a study of 1,464 badgers where the test was 93% specific and 49% sensitive against culture for M. bovis at necropsy. 8 Low sensitivity appears to be a common feature of serologic tests for M. bovis infection in many species, including badgers, 22 possums (Trichosurus vulpecula), 7 and cattle. 44 Detection of early stages of infection (before onset of clinical signs) is particularly difficult because of an inverse relationship between the courses of cell-mediated and humoral immunity, the latter being preferentially detected in cases of advanced disease.…”

Section: Discussionmentioning

confidence: 99%

Accuracy of Three Diagnostic Tests for Determining Mycobacterium Bovis Infection Status in Live-Sampled Wild Meerkats (Suricata Suricatta)

Drewe¹,

Dean

Michel

et al. 2009

J VET Diagn Invest

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Abstract. Accurate diagnosis of Mycobacterium bovis infection (bovine tuberculosis [bTB]) in live animals is notoriously problematic. The aim of the present study was to evaluate the diagnostic potential of 2 new serologic tests (multiantigen print immunoassay [MAPIA] and lateral flow immunoassay rapid test [RT]) in comparison with mycobacterial culture of tracheal washes for determining M. bovis infection status in a freeranging population of wild meerkats (Suricata suricatta). During a longitudinal study lasting 2.5 years, 240 individually identifiable meerkats were each sampled up to 8 times under anesthesia every 3 months. Diagnostic accuracy was determined through Bayesian and maximum likelihood estimations of sensitivity, specificity, and likelihood ratios for each diagnostic test when used independently and in parallel to classify the disease status of individual meerkats in the absence of a gold standard. Culture of tracheal washes was highly specific (0.99; 95% confidence interval [CI] 5 0.77, 1.00) but of low sensitivity (0.36; 95% CI 5 0.24, 0.50) for diagnosing M. bovis-infected individuals. The longitudinal nature of the study with repeated sampling of the same individual animals served simultaneously to improve chances of detecting infection and increase confidence in a negative result in individual animals repeatedly testing negative. Although MAPIA and RT were individually of limited diagnostic use, interpreting the results of these 2 tests in parallel produced estimates of sensitivity (0.83; 95% CI 5 0.67, 0.93) and specificity (0.73; 95% CI 5 0.62, 0.82) high enough to usefully inform decision making when determining exposure to bTB in wild meerkats and potentially other species in which bTB poses a diagnostic challenge.

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Section: Discussionmentioning

confidence: 99%

Accuracy of Three Diagnostic Tests for Determining Mycobacterium Bovis Infection Status in Live-Sampled Wild Meerkats (Suricata Suricatta)

Drewe¹,

Dean

Michel

et al. 2009

J VET Diagn Invest

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“…bovis expresses high levels of both MPB70 and MPB83 and these proteins are strongly recognized by the immune system during M. bovis infection in cattle and badgers (19,20). Although the proteins are expressed only at low levels in M. tuberculosis when grown in vitro, they are highly immunogenic during infection with live bacteria in mice (18) and man (21).…”

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confidence: 99%

The MPB83 Antigen from Mycobacterium bovis ContainsO-Linked Mannose and (1 → 3)-Mannobiose Moieties

Michell

Whelan

Wheeler

et al. 2003

Journal of Biological Chemistry

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Mycobacterium tuberculosis and Mycobacterium bovis, the causative agents of human and bovine tuberculosis, have been reported to express a range of surface and secreted glycoproteins, although only one of these has been subjected to detailed structural analysis. We describe the use of a genetic system, in conjunction with lectin binding, to characterize the points of attachment of carbohydrate moieties to the polypeptide backbone of a second mycobacterial glycoprotein, antigen MPB83 from M. bovis. Biochemical and structural analysis of the native MPB83 protein and derived peptides demonstrated the presence of 3 mannose units attached to two threonine residues. Mannose residues were joined by a (1 3 3) linkage, in contrast to the (1 3 2) linkage previously observed in antigen MPT32 from M. tuberculosis and the (1 3 2) and (1 3 6) linkages in other mycobacterial glycolipids and polysaccharides. The identification of glycosylated antigens within the M. tuberculosis complex raises the possibility that the carbohydrate moiety of these glycoproteins might be involved in pathogenesis, either by interaction with mannose receptors on host cells, or as targets or modulators of the cell-mediated immune response. Given such a possibility characterization of mycobacterial glycoproteins is a step toward understanding their functional role and elucidating the mechanisms of mycobacterial glycosylation.Protein glycosylation is ubiquitous in eukaryotes and the diverse array of carbohydrate moieties that can be fashioned to a polypeptide backbone makes glycoproteins ideal molecules to allow specific interactions with other molecules. In contrast, the number of reports of this post-translational modification by prokaryotes is comparatively low. Examples of eubacterial glycoproteins identified to date include cell surface or secreted proteins of pathogens that have antigenic properties and play a role in host pathogen interaction (1-6). Glycosylation of pilin has been postulated to enhance the adherence of Neisseria meningitidis to endothelial cells (2), whereas N-glycosylation of the platelet aggregation-associated protein of Staphylococcus sanguis may promote colonization of the endocardium (7, 8).Removal of a sero-specific glycosyl moiety from the flagellin of Campylobacter jejuni has been reported to alter the O antigenicity of the organism (9, 10). Similarly, Romain et al. (11) demonstrated that removal of covalently bound mannose from the Mycobacterium tuberculosis antigen MPT32 reduced by 10-fold its ability to elicit a delayed-type hypersensitivity reaction in guinea pigs immunized with Mycobacterium bovis BCG.M. tuberculosis and M. bovis are closely related members of the "M. tuberculosis complex" (MTb complex) that secrete a series of immunodominant antigens that have been reported to be glycosylated, on the basis of their ability to bind lectins such as concanavalin A (ConA) 1 (12-15). Structural confirmation of protein glycosylation by mycobacteria was provided by detailed chemical compositional analysis of MPT32, a 45...

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“…In vitro diagnostics that can be used to test live animals are required, for instance, to further develop vaccination strategies. Antibody-based assays have been developed (13,14), but they lack sensitivity (3,14), as they do for cattle (23). In comparison, assays based on measurement of cellular responses produce better results (6,23).…”

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confidence: 99%

Development and Evaluation of a Test for Tuberculosis in Live European Badgers (Meles meles) Based on Measurement of Gamma Interferon mRNA by Real-Time PCR

et al. 2007

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A real-time PCR assay for the measurement of gamma interferon (IFN-␥) mRNA in European badger (Meles meles) blood cultures was developed. The levels of IFN-␥ mRNA in blood cultures stimulated with either bovine or avian tuberculin or specific mycobacterial antigens were compared with those in a nonstimulated control blood culture as the basis for determining the tuberculosis (TB) status of live badgers. The assay was validated by testing 247 animals for which there were matching data from postmortem examination and culture of tissues. Relative changes in the levels of IFN-␥ mRNA in response to bovine tuberculin and specific antigens were found to be greater among badgers with tissues positive for TB on culture. The test was at its most accurate (87% of test results were correct) by using blood cultures containing bovine tuberculin as the antigen and when the response to avian tuberculin was taken into account by subtracting the avian tuberculin response from the bovine tuberculin response. At a specificity of 90.7%, the test was 70.6% sensitive. At the same specificity, the current serological enzyme-linked immunosorbent assay for TB in badgers was only 53% sensitive. This work demonstrates that measurement of IFN-␥ mRNA by real-time PCR is a valid method for the detection of TB in live badgers and may provide an alternative to the current serological methods of diagnosis, the Brock test. The testing procedure can be completed within 5 h of receipt of the blood culture samples. In addition, the use of a molecular biology-based test offers the potential to fully automate the testing procedure through the use of robotics.There has been a sharp rise in the incidence of bovine tuberculosis (TB) in cattle in the United Kingdom since the early 1990s (1). This adversely affects animal health and welfare and is a cause of considerable economic loss to farmers and the government. The European badger (Meles meles) has been implicated in the spread of disease by acting as a source of Mycobacterium bovis infection in cattle (9, 10). The accurate diagnosis of M. bovis infection in badgers can be achieved by bacteriological culture, but only for samples obtained postmortem. In vitro diagnostics that can be used to test live animals are required, for instance, to further develop vaccination strategies. Antibody-based assays have been developed (13, 14), but they lack sensitivity (3, 14), as they do for cattle (23). In comparison, assays based on measurement of cellular responses produce better results (6, 23). However, those developed to date for badgers are impractical for routine use (6). Tests analogous to that developed for the diagnosis of TB in cattle by the detection of gamma interferon (IFN-␥) in whole-blood cell cultures (21) have yet to be validated for badgers, although the badger IFN-␥ gene has been cloned and a polyclonal antiserum to the cytokine has been produced (7). Having cloned the badger IFN-␥ gene, we investigated the possibility of using IFN-␥ mRNA levels as a measure of the cell-mediated response to myco...

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Serodiagnosis of Mycobacterium bovis infection in badgers: development of an indirect ELISA using a 25 kDa antigen

Cited by 87 publications

References 0 publications

Accuracy of Three Diagnostic Tests for Determining Mycobacterium Bovis Infection Status in Live-Sampled Wild Meerkats (Suricata Suricatta)

Accuracy of Three Diagnostic Tests for Determining Mycobacterium Bovis Infection Status in Live-Sampled Wild Meerkats (Suricata Suricatta)

The MPB83 Antigen from Mycobacterium bovis ContainsO-Linked Mannose and (1 → 3)-Mannobiose Moieties

Development and Evaluation of a Test for Tuberculosis in Live European Badgers (Meles meles) Based on Measurement of Gamma Interferon mRNA by Real-Time PCR

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